Following spinal cord transection in the sea lamprey, axons regenerate selectively in their correct paths. Several molecules belonging to the semaphorin/collapsin and netrin families are thought to help guide axons during embryonic development and may also play a role in guiding regeneration in the injured, mature CNS. In previous studies, expression of netrin and semaphorin receptors occurred only in bad-regenerating neurons. Thus these guidance molecules may act as chemorepellents preventing axons from regenerating to their targets in the spinal cord. Aim 1. We will determine whether inhibiting expression of netrin and semaphorin receptors will enhance the regenerative capacities of identified reticulospinal neurons. Expression in these neurons will be inhibited by retrograde transport of fluoroscein-tagged antisense morpholino oligonucleotides injected at the site of spinal cord transection. The effect on distance and direction of regeneration will be determined by intra-axonal injection of tracer and compared with those in control animals injected with random oligonucleotides. Aim 2. Because chemorepulsion can be converted to attraction by manipulation of the cyclic guanosine-monophosphate(cGMP) and cyclic adenosine monophosphate (cAMP) signaling pathways, we will determine the effect of altering intracellular cAMP and/or cGMP concentrations by extracellular injections of membrane-permeable analogs on the regenerative capacities and/or pathway specificities of identified reticulo-spinal neurons. Aim 3. We will continue our search for axonal guidance molecules that are expessed preferentially in good-regenerating neurons. We will determine whether ephrins and their receptors Eph, the new semaphorin receptor CD72, intergins, and RTPR guide regenerating axons in their correct paths. Using primers from partial sequences already cloned and degenerate primers for others, we will clone by RTPCR the lamprey homologs of CD 72, ephrins, and their corresponding receptors, and determine the anatomical distribution of their mRNAs by semiquantitative in situ hybridization. The effect of spinal transection on their expression will be correlated in identified reticulospinal neurons with their regenerative capacities. By elucidating the mechanisms of regeneration in CNS axons, these experiments may lead to improved therapeutic approaches to inducing regeneration and functional recovery in human patients following spinal cord injury, head trauma or stroke.